A gene mutation is a permanent alteration in the DNA sequence that makes up a gene. Mutations can affect anywhere from a base pair to a large segment of a chromosome. Gene mutations can be classified in two major categories: (i) Germline or hereditary mutations are inherited from a parent and are present throughout a lifetime in virtually every cell in the body. Somatic or acquired mutations occur at some time periods during a lifetime and are present only in certain cells, not in every cell in the body. There are many different types of gene mutations, such as genetic code substitution, insertion, deletion or frameshift.
Cancers result from the accumulation of mutations in critical genes that alter normal programmes of cell proliferation, differentiation and death. Only about 5% to 10% of all cancers are thought to be related to germline mutations, and the rest are associated with somatic mutations. Fractions of neurodegenerative diseases such as Alzheimer disease (AD), Parkinson disease (PD), Huntington's disease (HD), and amyotrophic lateral sclerosis (ALS), are caused either by germline or somatic mutations (Kennedy et al., 2012).
Other common mutation-related disorders include, but not limited, 22q11.2 deletion syndrome, Angelman syndrome, Canavan disease, Charcot-Marie-Tooth disease, Color blindness, Cri du chat, Cystic fibrosis, Down syndrome, Duchenne muscular dystrophy, Haemochromatosis, Haemophilia, Klinefelter syndrome, Neurofibromatosis, Phenylketonuria, Polycystic kidney disease, Prader-Willi syndrome, Sickle-cell disease, Tay-Sachs disease, and Turner syndrome. The advances in molecular biology technologies have tremendously accelerated the discovery of causative genes. Despite this progress, however, the mutations causing a substantial number of diseases remain to be identified.
Mutations can occur at multiple sites of a same protein. For example, Factor XI mutation sites include: Met-18Ile, Ser-4Leu, Gly-1Arg, Asp16His, Val20Ala, Pro23Leu, Pro23Gln, Ser24Arg, Cys28Phe, Gln29His, Thr33Pro, Thr33Ile, Tyr35His, Cys38Arg, Cys38Trp, Pro48Leu, Pro52Leu, Arg54Pro, Thr57Ile, Cys58Arg, Cys58Phe, Cys58Tyr, Pro69Thr, Gly79Ala, Ser81Tyr, Lys83Arg, Gln88Stop, Cys92Gly, Met102Thr, Gly104Asp, Cys122Tyr, Thr123Met, Asp125Asp, His127Arg, Cys128Stop, Thr132Met, Tyr133Ser, Tyr133Cys, Ala134Pro, Arg144Cys, Gly155Glu, Leu172Pro, Ala181Val, Cys182Tyr, Arg184Gly, Pro188Ser, Asp198Asn, Cys212Ser, Cys212Arg, Phe221Ser, Ser225Phe, Glu226Arg, Trp228Cys, Arg234Lys, Arg234Ile, Arg234Ser, Cys237Tyr, Glu243Asp, Gly245Glu, Ser248Asn, Thr249Thr, Arg250Cys, Arg250His, Lys252Ile, Gly259Ser, Ile269Ile, Phe283Leu, Ile290Phe, Ile290Thr, Glu297Lys, Glu297Stop, Leu302Pro, Thr304Ile, Val307Phe, Arg308Cys, Cys309Stop, Thr313Ile, Cys321Phe, Glu323Lys, Gly336Arg, Ile341Met, Leu342Pro, Gly344Arg, Gly350Arg, Gly350Glu, Gly350Ala, Tyr351Ser, Tyr351 Stop, Leu355Ser, Cys356Arg, Val371Ile, Gly372Ala, Ala375Val, Arg378Cys, Gly379Gly, Trp381Leu, Trp381Arg, Pro382Leu, Trp383Stop, Thr386Asn, His388Pro, Thr389Pro, Thr390Pro, Cys398Tyr, Gly400Ser, Gly400Val, Ser401Ala, Gln406Stop, Trp407Cys, Thr410Ile, Ala412Ser, Ala412Thr, Ala412Val, Arg425Cys, Cys427Tyr, Ser428Gly, Gln433Glu, Phe442Val, Glu447Stop, Gly460Arg, Thr475Ile, Arg479Stop, Cys482Arg, Cys482Trp, Ser485Pro, Tyr493His, Trp497Cys, Val498Met, Trp501Stop, Trp501Cys, Lys518Asn, Pro520Leu, Cys527Tyr, Gly544Ser, Glu547Lys, Asp551Asp, Gly555Glu, Asp556Gly, Cys563Phe
Gene mutations may generate dysfunctional proteins, thus causing diseases. For example, the RAS-RAF-MEK-ERK-MAP kinase pathway regulates cellular growth. RAS mutation occurs in about 15% of human cancer. BRAF somatic missense mutations are found in 66% of malignant melanomas, among which a single substitution (V599E) accounts for 80%.
Gene mutations can be identified either by detecting mutated genes or the proteins encoded by mutated genes (referring to as mutant proteins hereafter). Some mutant proteins are disease-specific biomarkers, as identification of these mutant proteins is critical for disease diagnosis, staging, treatment, and prognosis. For that reason, mutation-specific antibodies are unmet needs for detecting mutation-related disease-specific biomarkers. In some cases, when a mutant protein is significantly different with the normal protein, an antibody may be generated to recognize specifically the mutant protein. However, a large number of disease-related mutant proteins are encoded by missense mutations, leading to only one amino acid substitutions in the mutant protein. For example, Wood et al. (2007) reported that the great majority of gene mutations are single-base substitutions (92.7%), with 81.9% resulting in missense changes. These subtle changes in mutant proteins make generating mutation-specific antibodies extremely difficult. For example, although v-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS) and tumor protein p53 (TP53) are two of the most commonly mutated and intensely studied cancer genes, there still are no antibodies that can reliably distinguish mutant from normal versions of these proteins (Wang et al., 2011). Therefore, novel method to generate mutation-specific antibodies is an unmet need for identifying mutant proteins.